Bidirectional modulation of TCA cycle metabolites and anaplerosis by metformin and its combination with SGLT2i.

BACKGROUND: Metformin and sodium-glucose-cotransporter-2 inhibitors (SGLT2i) are cornerstone therapies for managing hyperglycemia in diabetes. However, their detailed impacts on metabolic processes, particularly within the citric acid (TCA) cycle and its anaplerotic pathways, remain unclear. This study investigates the tissue-specific metabolic effects of metformin, both as a monotherapy and in combination with SGLT2i, on the TCA cycle and associated anaplerotic reactions in both mice and humans. METHODS: Metformin-specific metabolic changes were initially identified by comparing metformin-treated diabetic mice (MET) with vehicle-treated db/db mice (VG). These findings were then assessed in two human cohorts (KORA and QBB) and a longitudinal KORA study of metformin-naïve patients with Type 2 Diabetes (T2D). We also compared MET with db/db mice on combination therapy (SGLT2i + MET). Metabolic profiling analyzed 716 metabolites from plasma, liver, and kidney tissues post-treatment, using linear regression and Bonferroni correction for statistical analysis, complemented by pathway analyses to explore the pathophysiological implications. RESULTS: Metformin monotherapy significantly upregulated TCA cycle intermediates such as malate, fumarate, and α-ketoglutarate (α-KG) in plasma, and anaplerotic substrates including hepatic glutamate and renal 2-hydroxyglutarate (2-HG) in diabetic mice. Downregulated hepatic taurine was also observed. The addition of SGLT2i, however, reversed these effects, such as downregulating circulating malate and α-KG, and hepatic glutamate and renal 2-HG, but upregulated hepatic taurine. In human T2D patients on metformin therapy, significant systemic alterations in metabolites were observed, including increased malate but decreased citrulline. The bidirectional modulation of TCA cycle intermediates in mice influenced key anaplerotic pathways linked to glutaminolysis, tumorigenesis, immune regulation, and antioxidative responses. CONCLUSION: This study elucidates the specific metabolic consequences of metformin and SGLT2i on the TCA cycle, reflecting potential impacts on the immune system. Metformin shows promise for its anti-inflammatory properties, while the addition of SGLT2i may provide liver protection in conditions like metabolic dysfunction-associated steatotic liver disease (MASLD). These observations underscore the importance of personalized treatment strategies.

Inhibition of mammalian mtDNA transcription acts paradoxically to reverse diet-induced hepatosteatosis and obesity.

The oxidative phosphorylation system1 in mammalian mitochondria plays a key role in transducing energy from ingested nutrients2. Mitochondrial metabolism is dynamic and can be reprogrammed to support both catabolic and anabolic reactions, depending on physiological demands or disease states. Rewiring of mitochondrial metabolism is intricately linked to metabolic diseases and promotes tumour growth3-5. Here, we demonstrate that oral treatment with an inhibitor of mitochondrial transcription (IMT)6 shifts whole-animal metabolism towards fatty acid oxidation, which, in turn, leads to rapid normalization of body weight, reversal of hepatosteatosis and restoration of normal glucose tolerance in male mice on a high-fat diet. Paradoxically, the IMT treatment causes a severe reduction of oxidative phosphorylation capacity concomitant with marked upregulation of fatty acid oxidation in the liver, as determined by proteomics and metabolomics analyses. The IMT treatment leads to a marked reduction of complex I, the main dehydrogenase feeding electrons into the ubiquinone (Q) pool, whereas the levels of electron transfer flavoprotein dehydrogenase and other dehydrogenases connected to the Q pool are increased. This rewiring of metabolism caused by reduced mtDNA expression in the liver provides a principle for drug treatment of obesity and obesity-related pathology.

Novel markers and networks related to restored skeletal muscle transcriptome after bariatric surgery.

OBJECTIVE: The aim of this study was to discover novel markers underlying the improvement of skeletal muscle metabolism after bariatric surgery. METHODS: Skeletal muscle transcriptome data of lean people and people with obesity, before and 1 year after bariatric surgery, were subjected to weighted gene co-expression network analysis (WGCNA) and least absolute shrinkage and selection operator (LASSO) regression. Results of LASSO were confirmed in a replication cohort. RESULTS: The expression levels of 440 genes differing between individuals with and without obesity were no longer different 1 year after surgery, indicating restoration. WGCNA clustered 116 genes with normalized expression in one major module, particularly correlating to weight loss and decreased plasma free fatty acids (FFA), 44 of which showed an obesity-related phenotype upon deletion in mice. Among the genes of the major module, 105 represented prominent markers for reduced FFA concentration, including 55 marker genes for decreased BMI in both the discovery and replication cohorts. CONCLUSIONS: Previously unknown gene networks and marker genes underlined the important role of FFA in restoring muscle gene expression after bariatric surgery and further suggest novel therapeutic targets for obesity.

Knockout mice are an important tool for human monogenic heart disease studies.

Mouse models are relevant to studying the functionality of genes involved in human diseases; however, translation of phenotypes can be challenging. Here, we investigated genes related to monogenic forms of cardiovascular disease based on the Genomics England PanelApp and aligned them to International Mouse Phenotyping Consortium (IMPC) data. We found 153 genes associated with cardiomyopathy, cardiac arrhythmias or congenital heart disease in humans, of which 151 have one-to-one mouse orthologues. For 37.7% (57/151), viability and heart data captured by electrocardiography, transthoracic echocardiography, morphology and pathology from embryos and young adult mice are available. In knockout mice, 75.4% (43/57) of these genes showed non-viable phenotypes, whereas records of prenatal, neonatal or infant death in humans were found for 35.1% (20/57). Multisystem phenotypes are common, with 58.8% (20/34) of heterozygous (homozygous lethal) and 78.6% (11/14) of homozygous (viable) mice showing cardiovascular, metabolic/homeostasis, musculoskeletal, hematopoietic, nervous system and/or growth abnormalities mimicking the clinical manifestations observed in patients. These IMPC data are critical beyond cardiac diagnostics given their multisystemic nature, allowing detection of abnormalities across physiological systems and providing a valuable resource to understand pleiotropic effects.

Monitoring longitudinal disease progression in a novel murine Kit tumor model using high-field MRI.

Animal models are an indispensable platform used in various research disciplines, enabling, for example, studies of basic biological mechanisms, pathological processes and new therapeutic interventions. In this study, we applied magnetic resonance imaging (MRI) to characterize the clinical picture of a novel N-ethyl-N-nitrosourea-induced Kit-mutant mouse in vivo. Seven C3H KitN824K/WT mutant animals each of both sexes and their littermates were monitored every other month for a period of twelve months. MRI relaxometry data of hematopoietic bone marrow and splenic tissue as well as high-resolution images of the gastrointestinal organs were acquired. Compared with controls, the mutants showed a dynamic change in the shape and volume of the cecum and enlarged Peyer´s patches were identified throughout the entire study. Mammary tumors were observed in the majority of mutant females and were first detected at eight months of age. Using relaxation measurements, a substantial decrease in longitudinal relaxation times in hematopoietic tissue was detected in mutants at one year of age. In contrast, transverse relaxation time of splenic tissue showed no differences between genotypes, except in two mutant mice, one of which had leukemia and the other hemangioma. In this study, in vivo MRI was used for the first time to thoroughly characterize the evolution of systemic manifestations of a novel Kit-induced tumor model and to document the observable organ-specific disease cascade.

Mass spectrometry-based draft of the mouse proteome.

The laboratory mouse ranks among the most important experimental systems for biomedical research and molecular reference maps of such models are essential informational tools. Here, we present a quantitative draft of the mouse proteome and phosphoproteome constructed from 41 healthy tissues and several lines of analyses exemplify which insights can be gleaned from the data. For instance, tissue- and cell-type resolved profiles provide protein evidence for the expression of 17,000 genes, thousands of isoforms and 50,000 phosphorylation sites in vivo. Proteogenomic comparison of mouse, human and Arabidopsis reveal common and distinct mechanisms of gene expression regulation and, despite many similarities, numerous differentially abundant orthologs that likely serve species-specific functions. We leverage the mouse proteome by integrating phenotypic drug (n > 400) and radiation response data with the proteomes of 66 pancreatic ductal adenocarcinoma (PDAC) cell lines to reveal molecular markers for sensitivity and resistance. This unique atlas complements other molecular resources for the mouse and can be explored online via ProteomicsDB and PACiFIC.

Does a Hypertrophying Muscle Fibre Reprogramme its Metabolism Similar to a Cancer Cell?

In 1924, Otto Warburg asked "How does the metabolism of a growing tissue differ from that of a non-growing tissue?" Currently, we know that proliferating healthy and cancer cells reprogramme their metabolism. This typically includes increased glucose uptake, glycolytic flux and lactate synthesis. A key function of this reprogramming is to channel glycolytic intermediates and other metabolites into anabolic reactions such as nucleotide-RNA/DNA synthesis, amino acid-protein synthesis and the synthesis of, for example, acetyl and methyl groups for epigenetic modification. In this review, we discuss evidence that a hypertrophying muscle similarly takes up more glucose and reprogrammes its metabolism to channel energy metabolites into anabolic pathways. We specifically discuss the functions of the cancer-associated enzymes phosphoglycerate dehydrogenase and pyruvate kinase muscle 2 in skeletal muscle. In addition, we ask whether increased glucose uptake by a hypertrophying muscle explains why muscularity is often negatively associated with type 2 diabetes mellitus and obesity.

On the Nature of Murine Radiation-Induced Subcapsular Cataracts: Optical Coherence Tomography-Based Fine Classification, In Vivo Dynamics and Impact on Visual Acuity.

Ionizing radiation is widely known to induce various kinds of lens cataracts, of which posterior subcapsular cataracts (PSCs) have the highest prevalence. Despite some studies regarding the epidemiology and biology of radiation-induced PSCs, the mechanism underscoring the formation of this type of lesions and their dose dependency remain uncertain. Within the current study, our team investigated the in vivo characteristics of PSCs in B6C3F1 mice (F1-hybrids of BL6 × C3H) that received 0.5-2 Gy γ-ray irradiation after postnatal day 70. For purposes of assessing lenticular damages, spectral domain optical coherence tomography was utilized, and the visual acuity of the mice was measured to analyze their levels of visual impairment, and histological sections were then prepared in to characterize in vivo phenotypes. Three varying in vivo phenotype anterior and posterior lesions were thus revealed and correlated with the applied doses to understand their marginal influence on the visual acuity of the studied mice. Histological data indicated no significantly increased odds ratios for PSCs below a dose of 1 Gy at the end of the observation time. Furthermore, our team demonstrated that when the frequencies of the posterior and anterior lesions were calculated at early time points, their responses were in accordance with a deterministic model, whereas at later time points, their responses were better described via a stochastic model. The current study will aid in honing the current understanding of radiation-induced cataract formation and contributes greatly to addressing the fundamental questions of lens dose response within the field of radiation biology.

Creld1 regulates myocardial development and function.

CRELD1 (Cysteine-Rich with EGF-Like Domains 1) is a risk gene for non-syndromic atrioventricular septal defects in human patients. In a mouse model, Creld1 has been shown to be essential for heart development, particularly in septum and valve formation. However, due to the embryonic lethality of global Creld1 knockout (KO) mice, its cell type-specific function during peri- and postnatal stages remains unknown. Here, we generated conditional Creld1 KO mice lacking Creld1 either in the endocardium (KOTie2) or the myocardium (KOMyHC). Using a combination of cardiac phenotyping, histology, immunohistochemistry, RNA-sequencing, and flow cytometry, we demonstrate that Creld1 function in the endocardium is dispensable for heart development. Lack of myocardial Creld1 causes extracellular matrix remodeling and trabeculation defects by modulation of the Notch1 signaling pathway. Hence, KOMyHC mice die early postnatally due to myocardial hypoplasia. Our results reveal that Creld1 not only controls the formation of septa and valves at an early stage during heart development, but also cardiac maturation and function at a later stage. These findings underline the central role of Creld1 in mammalian heart development and function.

Dose-dependent long-term effects of a single radiation event on behaviour and glial cells.

PURPOSE: The increasing use of low-dose ionizing radiation in medicine requires a systematic study of its long-term effects on the brain, behaviour and its possible association with neurodegenerative disease vulnerability. Therefore, we analysed the long-term effects of a single low-dose irradiation exposure at 10 weeks of age compared to medium and higher doses on locomotor, emotion-related and sensorimotor behaviour in mice as well as on hippocampal glial cell populations. MATERIALS AND METHODS: We determined the influence of radiation dose (0, 0.063, 0.125 or 0.5 Gy), time post-irradiation (4, 12 and 18 months p.i.), sex and genotype (wild type versus mice with Ercc2 DNA repair gene point mutation) on behaviour. RESULTS: The high dose (0.5 Gy) had early-onset adverse effects at 4 months p.i. on sensorimotor recruitment and late-onset negative locomotor effects at 12 and 18 months p.i. Notably, the low dose (0.063 Gy) produced no early effects but subtle late-onset (18 months) protective effects on sensorimotor recruitment and exploratory behaviour. Quantification and morphological characterization of the microglial and the astrocytic cells of the dentate gyrus 24 months p.i. indicated heightened immune activity after high dose irradiation (0.125 and 0.5 Gy) while conversely, low dose (0.063 Gy) induced more neuroprotective features. CONCLUSION: This is one of the first studies demonstrating such long-term and late-onset effects on brain and behaviour after a single radiation event in adulthood.

Inhibition of LTßR signalling activates WNT-induced regeneration in lung.

Lymphotoxin ß-receptor (LTßR) signalling promotes lymphoid neogenesis and the development of tertiary lymphoid structures1,2, which are associated with severe chronic inflammatory diseases that span several organ systems3-6. How LTßR signalling drives chronic tissue damage particularly in the lung, the mechanism(s) that regulate this process, and whether LTßR blockade might be of therapeutic value have remained unclear. Here we demonstrate increased expression of LTßR ligands in adaptive and innate immune cells, enhanced non-canonical NF-κB signalling, and enriched LTßR target gene expression in lung epithelial cells from patients with smoking-associated chronic obstructive pulmonary disease (COPD) and from mice chronically exposed to cigarette smoke. Therapeutic inhibition of LTßR signalling in young and aged mice disrupted smoking-related inducible bronchus-associated lymphoid tissue, induced regeneration of lung tissue, and reverted airway fibrosis and systemic muscle wasting. Mechanistically, blockade of LTßR signalling dampened epithelial non-canonical activation of NF-κB, reduced TGFß signalling in airways, and induced regeneration by preventing epithelial cell death and activating WNT/ß-catenin signalling in alveolar epithelial progenitor cells. These findings suggest that inhibition of LTßR signalling represents a viable therapeutic option that combines prevention of tertiary lymphoid structures1 and inhibition of apoptosis with tissue-regenerative strategies.

Characterization of Hormone-Dependent Pathways in Six Human Prostate-Cancer Cell Lines: A Gene-Expression Study.

Prostate cancer (PCa), the most incident cancer in men, is tightly regulated by endocrine signals. A number of different PCa cell lines are commonly used for in vitro experiments, but these are of diverse origin, and have very different cell-proliferation rates and hormone-response capacities. By analyzing the gene-expression pattern of main hormone pathways, we systematically compared six PCa cell lines and parental primary cells. We compared these cell lines (i) with each other and (ii) with PCa tissue samples from 11 patients. We found major differences in the gene-expression levels of androgen, insulin, estrogen, and oxysterol signaling between PCa tissue and cell lines, and between different cell lines. Our systematic characterization gives researchers a solid basis to choose the appropriate PCa cell model for the hormone pathway of interest.

Transcript Levels of Aldo-Keto Reductase Family 1 Subfamily C (AKR1C) Are Increased in Prostate Tissue of Patients with Type 2 Diabetes.

Aldo-keto reductase family 1 (AKR1) enzymes play a crucial role in diabetic complications. Since type 2 diabetes (T2D) is associated with cancer progression, we investigated the impact of diabetes on AKR1 gene expression in the context of prostate cancer (PCa) development. In this study, we analyzed benign (BEN) prostate and PCa tissue of patients with and without T2D. Furthermore, to replicate hyperglycemia in vitro, we treated the prostate adenocarcinoma cell line PC3 with increasing glucose concentrations. Gene expression was quantified using real-time qPCR. In the prostate tissue of patients with T2D, AKR1C1 and AKR1C2 transcripts were higher compared to samples of patients without diabetes. In PC3 cells, high glucose treatment induced the gene expression levels of AKR1C1, C2, and C3. Furthermore, both in human tissue and in PC3 cells, the transcript levels of AKR1C1, C2, and C3 showed positive associations with oncogenes, which are involved in proliferation processes and HIF1α and NFκB pathways. These results indicate that in the prostate glands of patients with T2D, hyperglycemia could play a pivotal role by inducing the expression of AKR1C1, C2, and C3. The higher transcript level of AKR1C was furthermore associated with upregulated HIF1α and NFκB pathways, which are major drivers of PCa carcinogenesis.

METTL6 is a tRNA m3C methyltransferase that regulates pluripotency and tumor cell growth.

Recently, covalent modifications of RNA, such as methylation, have emerged as key regulators of all aspects of RNA biology and have been implicated in numerous diseases, for instance, cancer. Here, we undertook a combination of in vitro and in vivo screens to test 78 potential methyltransferases for their roles in hepatocellular carcinoma (HCC) cell proliferation. We identified methyltransferase-like protein 6 (METTL6) as a crucial regulator of tumor cell growth. We show that METTL6 is a bona fide transfer RNA (tRNA) methyltransferase, catalyzing the formation of 3-methylcytidine at C32 of specific serine tRNA isoacceptors. Deletion of Mettl6 in mouse stem cells results in changes in ribosome occupancy and RNA levels, as well as impaired pluripotency. In mice, Mettl6 knockout results in reduced energy expenditure. We reveal a previously unknown pathway in the maintenance of translation efficiency with a role in maintaining stem cell self-renewal, as well as impacting tumor cell growth profoundly.

Physiological relevance of the neuronal isoform of inositol-1,4,5-trisphosphate 3-kinases in mice.

Inositol-1,4,5-trisphosphate 3-kinase-A (ITPKA) is the neuronal isoform of ITPKs and exhibits both actin bundling and InsP3kinase activity. In addition to neurons, ITPKA is ectopically expressed in tumor cells, where its oncogenic activity increases tumor cell malignancy. In order to analyze the physiological relevance of ITPKA, here we performed a broad phenotypic screening of itpka deficient mice. Our data show that among the neurobehavioral tests analyzed, itpka deficient mice reacted faster to a hotplate, prepulse inhibition was impaired and the accelerating rotarod test showed decreased latency of itpka deficient mice to fall. These data indicate that ITPKA is involved in the regulation of nociceptive pathways, sensorimotor gating and motor learning. Analysis of extracerebral functions in control and itpka deficient mice revealed significantly reduced glucose, lactate, and triglyceride plasma concentrations in itpka deficient mice. Based on this finding, expression of ITPKA was analyzed in extracerebral tissues and the highest level was found in the small intestine. However, functional studies on CaCo-2 control and ITPKA depleted cells showed that glucose, as well as triglyceride uptake, were not significantly different between the cell lines. Altogether, these data show that ITPKA exhibits distinct functions in the central nervous system and reveal an involvement of ITPKA in energy metabolism.

Irp2 regulates insulin production through iron-mediated Cdkal1-catalyzed tRNA modification.

Regulation of cellular iron homeostasis is crucial as both iron excess and deficiency cause hematological and neurodegenerative diseases. Here we show that mice lacking iron-regulatory protein 2 (Irp2), a regulator of cellular iron homeostasis, develop diabetes. Irp2 post-transcriptionally regulates the iron-uptake protein transferrin receptor 1 (TfR1) and the iron-storage protein ferritin, and dysregulation of these proteins due to Irp2 loss causes functional iron deficiency in ß cells. This impairs Fe-S cluster biosynthesis, reducing the function of Cdkal1, an Fe-S cluster enzyme that catalyzes methylthiolation of t6A37 in tRNALysUUU to ms2t6A37. As a consequence, lysine codons in proinsulin are misread and proinsulin processing is impaired, reducing insulin content and secretion. Iron normalizes ms2t6A37 and proinsulin lysine incorporation, restoring insulin content and secretion in Irp2-/- ß cells. These studies reveal a previously unidentified link between insulin processing and cellular iron deficiency that may have relevance to type 2 diabetes in humans.

CRN2 binds to TIMP4 and MMP14 and promotes perivascular invasion of glioblastoma cells.

CRN2 is an actin filament binding protein involved in the regulation of various cellular processes including cell migration and invasion. CRN2 has been implicated in the malignant progression of different types of human cancer. We used CRN2 knock-out mice for analyses as well as for crossbreeding with a Tp53/Pten knock-out glioblastoma mouse model. CRN2 knock-out mice were subjected to a phenotyping screen at the German Mouse Clinic. Murine glioblastoma tissue specimens as well as cultured murine brain slices and glioblastoma cell lines were investigated by immunohistochemistry, immunofluorescence, and cell biological experiments. Protein interactions were studied by immunoprecipitation, pull-down, and enzyme activity assays. CRN2 knock-out mice displayed neurological and behavioural alterations, e.g. reduced hearing sensitivity, reduced acoustic startle response, hypoactivity, and less frequent urination. While glioblastoma mice with or without the additional CRN2 knock-out allele exhibited no significant difference in their survival rates, the increased levels of CRN2 in transplanted glioblastoma cells caused a higher tumour cell encasement of murine brain slice capillaries. We identified two important factors of the tumour microenvironment, the tissue inhibitor of matrix metalloproteinase 4 (TIMP4) and the matrix metalloproteinase 14 (MMP14, synonym: MT1-MMP), as novel binding partners of CRN2. All three proteins mutually interacted and co-localised at the front of lamellipodia, and CRN2 was newly detected in exosomes. On the functional level, we demonstrate that CRN2 increased the secretion of TIMP4 as well as the catalytic activity of MMP14. Our results imply that CRN2 represents a pro-invasive effector within the tumour cell microenvironment of glioblastoma multiforme.

Towards frailty biomarkers: Candidates from genes and pathways regulated in aging and age-related diseases.

OBJECTIVE: Use of the frailty index to measure an accumulation of deficits has been proven a valuable method for identifying elderly people at risk for increased vulnerability, disease, injury, and mortality. However, complementary molecular frailty biomarkers or ideally biomarker panels have not yet been identified. We conducted a systematic search to identify biomarker candidates for a frailty biomarker panel. METHODS: Gene expression databases were searched (http://genomics.senescence.info/genes including GenAge, AnAge, LongevityMap, CellAge, DrugAge, Digital Aging Atlas) to identify genes regulated in aging, longevity, and age-related diseases with a focus on secreted factors or molecules detectable in body fluids as potential frailty biomarkers. Factors broadly expressed, related to several "hallmark of aging" pathways as well as used or predicted as biomarkers in other disease settings, particularly age-related pathologies, were identified. This set of biomarkers was further expanded according to the expertise and experience of the authors. In the next step, biomarkers were assigned to six "hallmark of aging" pathways, namely (1) inflammation, (2) mitochondria and apoptosis, (3) calcium homeostasis, (4) fibrosis, (5) NMJ (neuromuscular junction) and neurons, (6) cytoskeleton and hormones, or (7) other principles and an extensive literature search was performed for each candidate to explore their potential and priority as frailty biomarkers. RESULTS: A total of 44 markers were evaluated in the seven categories listed above, and 19 were awarded a high priority score, 22 identified as medium priority and three were low priority. In each category high and medium priority markers were identified. CONCLUSION: Biomarker panels for frailty would be of high value and better than single markers. Based on our search we would propose a core panel of frailty biomarkers consisting of (1) CXCL10 (C-X-C motif chemokine ligand 10), IL-6 (interleukin 6), CX3CL1 (C-X3-C motif chemokine ligand 1), (2) GDF15 (growth differentiation factor 15), FNDC5 (fibronectin type III domain containing 5), vimentin (VIM), (3) regucalcin (RGN/SMP30), calreticulin, (4) PLAU (plasminogen activator, urokinase), AGT (angiotensinogen), (5) BDNF (brain derived neurotrophic factor), progranulin (PGRN), (6) α-klotho (KL), FGF23 (fibroblast growth factor 23), FGF21, leptin (LEP), (7) miRNA (micro Ribonucleic acid) panel (to be further defined), AHCY (adenosylhomocysteinase) and KRT18 (keratin 18). An expanded panel would also include (1) pentraxin (PTX3), sVCAM/ICAM (soluble vascular cell adhesion molecule 1/Intercellular adhesion molecule 1), defensin α, (2) APP (amyloid beta precursor protein), LDH (lactate dehydrogenase), (3) S100B (S100 calcium binding protein B), (4) TGFß (transforming growth factor beta), PAI-1 (plasminogen activator inhibitor 1), TGM2 (transglutaminase 2), (5) sRAGE (soluble receptor for advanced glycosylation end products), HMGB1 (high mobility group box 1), C3/C1Q (complement factor 3/1Q), ST2 (Interleukin 1 receptor like 1), agrin (AGRN), (6) IGF-1 (insulin-like growth factor 1), resistin (RETN), adiponectin (ADIPOQ), ghrelin (GHRL), growth hormone (GH), (7) microparticle panel (to be further defined), GpnmB (glycoprotein nonmetastatic melanoma protein B) and lactoferrin (LTF). We believe that these predicted panels need to be experimentally explored in animal models and frail cohorts in order to ascertain their diagnostic, prognostic and therapeutic potential.

Disease model discovery from 3,328 gene knockouts by The International Mouse Phenotyping Consortium.

Although next-generation sequencing has revolutionized the ability to associate variants with human diseases, diagnostic rates and development of new therapies are still limited by a lack of knowledge of the functions and pathobiological mechanisms of most genes. To address this challenge, the International Mouse Phenotyping Consortium is creating a genome- and phenome-wide catalog of gene function by characterizing new knockout-mouse strains across diverse biological systems through a broad set of standardized phenotyping tests. All mice will be readily available to the biomedical community. Analyzing the first 3,328 genes identified models for 360 diseases, including the first models, to our knowledge, for type C Bernard-Soulier, Bardet-Biedl-5 and Gordon Holmes syndromes. 90% of our phenotype annotations were novel, providing functional evidence for 1,092 genes and candidates in genetically uncharacterized diseases including arrhythmogenic right ventricular dysplasia 3. Finally, we describe our role in variant functional validation with The 100,000 Genomes Project and others.

Extensive phenotypic characterization of a new transgenic mouse reveals pleiotropic perturbations in physiology due to mesenchymal hGH minigene expression.

The human growth hormone (hGH) minigene used for transgene stabilization in mice has been recently identified to be locally expressed in the tissues where transgenes are active and associated with phenotypic alterations. Here we extend these findings by analyzing the effect of the hGH minigene in TgC6hp55 transgenic mice which express the human TNFR1 under the control of the mesenchymal cell-specific CollagenVI promoter. These mice displayed a fully penetrant phenotype characterized by growth enhancement accompanied by perturbations in metabolic, skeletal, histological and other physiological parameters. Notably, this phenotype was independent of TNF-TNFR1 signaling since the genetic ablation of either Tnf or Tradd did not rescue the phenotype. Further analyses showed that the hGH minigene was expressed in several tissues, also leading to increased hGH protein levels in the serum. Pharmacological blockade of GH signaling prevented the development of the phenotype. Our results indicate that the unplanned expression of the hGH minigene in CollagenVI expressing mesenchymal cells can lead through local and/or systemic mechanisms to enhanced somatic growth followed by a plethora of primary and/or secondary effects such as hyperphagia, hypermetabolism, disturbed glucose homeostasis, altered hematological parameters, increased bone formation and lipid accumulation in metabolically critical tissues.